Chytré nanonástroje

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Tuning the surface coating of IONs toward efficient sonochemical tethering and sustained liberation of topoisomerase II poisons
(Dove Medical Press, 2019-09-17) Michálková, Hana; Strmiska, Vladislav; Kudr, Jiří; Škubalová, Zuzana; Tesařová, Barbora; Švec, Pavel; Richtera, Lukáš; Zítka, Ondřej; Adam, Vojtěch; Heger, Zbyněk
Background: Iron oxide nanoparticles (IONs) have been increasingly utilized in a wide spectrum of biomedical applications. Surface coatings of IONs can bestow a number of exceptional properties, including enhanced stability of IONs, increased loading of drugs or their controlled release. Methods: Using two-step sonochemical protocol, IONs were surface-coated with polyoxyethylene stearate, polyvinylpyrrolidone or chitosan for a loading of two distinct topo II poisons (doxorubicin and ellipticine). The cytotoxic behavior was tested in vitro against breast cancer (MDA-MB-231) and healthy epithelial cells (HEK-293 and HBL-100). In addition, biocompatibility studies (hemotoxicity, protein corona formation, binding of third complement component) were performed. Results: Notably, despite surface-coated IONs exhibited only negligible cytotoxicity, upon tethering with topo II poisons, synergistic or additional enhancement of cytotoxicity was found in MDA-MB-231 cells. Pronounced anti-migratory activity, DNA fragmentation, decrease in expression of procaspase-3 and enhancement of p53 expression were further identified upon exposure to surface-coated IONs with tethered doxorubicin and ellipticine. Moreover, surface-coated IONs nanoformulations of topo II poisons exhibited exceptional stability in human plasma with no protein corona and complement 3 binding, and only a mild induction of hemolysis in human red blood cells. Conclusion: The results imply a high potential of an efficient ultrasound-mediated surface functionalization of IONs as delivery vehicles to improve therapeutic efficiency of topo II poisons.
CdSe QD biosynthesis in yeast using tryptone-enriched media and their conjugation with a peptide hecate for bacterial detection and killing
(MDPI, 2019-10-31) Sur, Vishma Pratap; Komínková, Markéta; Buchtová, Žaneta; Doleželíková, Kristýna; Zítka, Ondřej; Moulick, Amitava
The physical and chemical synthesis methods of quantum dots (QDs) are generally unfavorable for biological applications. To overcome this limitation, the development of a novel "green" route to produce highly-fluorescent CdSe QDs constitutes a promising substitute approach. In the present work, CdSe QDs were biosynthesized in yeast Saccharomyces cerevisiae using a novel method, where we showed for the first time that the concentration of tryptone highly affects the synthesis process. The optimum concentration of tryptone was found to be 25 g/L for the highest yield. Different methods were used to optimize the QD extraction from yeast, and the best method was found to be by denaturation at 80 degrees C along with an ultrasound needle. Multiple physical characterizations including transmission electron microscopy (TEM), dynamic light scattering (DLS), energy-dispersive X-ray spectroscopy (EDX), and spectrophotometry confirmed the optical features size and shape distribution of the QDs. We showed that the novel conjugate of the CdSe QDs and a cell-penetrating peptide (hecate) can detect bacterial cells very efficiently under a fluorescent microscope. The conjugate also showed strong antibacterial activity against vancomycin-resistant Staphylococcus aureus (VRSA), methicillin-resistant Staphylococcus aureus (MRSA), and Escherichia coli, which may help us to cope with the problem of rising antibiotic resistance.
In Vitro Interactions between 17 beta-Estradiol and DNA Result in Formation of the Hormone-DNA Complexes
(MDPI, 2014-08-01) Heger, Zbyněk; Guráň, Roman; Zítka, Ondřej; Beklová, Miroslava; Adam, Vojtěch; Kizek, René
Beyond the role of 17 beta-estradiol (E-2) in reproduction and during the menstrual cycle, it has been shown to modulate numerous physiological processes such as cell proliferation, apoptosis, inflammation and ion transport in many tissues. The pathways in which estrogens affect an organism have been partially described, although many questions still exist regarding estrogens' interaction with biomacromolecules. Hence, the present study showed the interaction of four oligonucleotides (17, 20, 24 and/or 38-mer) with E-2. The strength of these interactions was evaluated using optical methods, showing that the interaction is influenced by three major factors, namely: oligonucleotide length, E-2 concentration and interaction time. In addition, the denaturation phenomenon of DNA revealed that the binding of E-2 leads to destabilization of hydrogen bonds between the nitrogenous bases of DNA strands resulting in a decrease of their melting temperatures (T-m). To obtain a more detailed insight into these interactions, MALDI-TOF mass spectrometry was employed. This study revealed that E-2 with DNA forms non-covalent physical complexes, observed as the mass shifts for app. 270 Da (Mr of E-2) to higher molecular masses. Taken together, our results indicate that E-2 can affect biomacromolecules, as circulating oligonucleotides, which can trigger mutations, leading to various unwanted effects.
Lead Ions Encapsulated in Liposomes and Their Effect on Staphylococcus aureus
(MDPI, 2013-12-01) Kenšová, Renáta; Blažková, Iva; Konečná, Marie; Kopel, Pavel; chudobova, Dagmar; Zítka, Ondřej; Vaculovičová, Markéta; Hynek, David; Adam, Vojtěch; Beklová, Miroslava; Kizek, René
The aim of the study was the preparation of a liposome complex with encapsulated lead ions, which were electrochemically detected. In particular, experiments were focused on the potential of using an electrochemical method for the determination of free and liposome-encapsulated lead and determination of the encapsulation efficiency preventing the lead toxicity. Primarily, encapsulation of lead ions in liposomes and confirmation of successful encapsulation by electrochemical methods was done. Further, the reduction effect of the liposome matrix on the detected electrochemical signal was monitored. Besides encapsulation itself, comparison of toxicity of free lead ions and lead ions encapsulated in liposome was tested. The calculated IC50 values for evaluating the lead cytotoxicity showed significant differences between the lead enclosed in liposomes (28 mu M) and free lead ions (237 mu M). From the cytotoxicity studies on the bacterial strain of S. aureus it was observed that the free lead ions are less toxic in comparison with lead encapsulated in liposomes. Liposomes appear to be a suitable carrier of various substances through the inner cavity. Due to the liposome structure the lead enclosed in the liposome is more easily accepted into the cell structure and the toxicity of the enclosed lead is higher in comparison to free lead ions.
A simple electrochemical biosensor for the detection of methylated DNA and for methyltransferase activity monitoring
(Mendel University in Brno, 2019-12-31) Dostálová, Eliška; Smolíková, Vendula; Birgusová, Eliška; Bytešníková, Zuzana; Richtera, Lukáš; Adam, Vojtěch
DNA methylation is one of the well-known epigenetic mechanism which plays a crucial role in the development of various diseases, such as cancer, cardiovascular diseases or diabetes. Electrochemical biosensors promise excellent results for clinical diagnostics, especially in term of their sensitivity, stability, selectivity, portability, they are cost-effective, easy-to-use and provides a fast response. In this study, simply, reliably and selective DNA based electrochemical biosensor for detection of methylated DNA was fabricated. The proposed biosensor was modified with the synthesised reduced graphene oxide combined with gold nanoparticles (rGO-AuNPs). This nanocomposite has shown a strong affinity to the DNA probe and demonstrated promising analytical characteristics. The electrochemical impedance spectroscopy (EIS) was used for the characterization of interface properties of the gold electrode (GE). Additionally, the sensitivity of the developed biosensor was performed by differential pulse voltammetry (DPV) to investigate the activity of enzyme methyltransferase M.SssI (MTase). Fabricated biosensor offers quite a low detection limit (LOD), which was 3.2 U/ml and limit of quantification (LOQ) was 3.3 U/ml.

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